Project 4 Abstract Human health is at risk due to environmental exposures to a wide range of chemical toxicants and endocrine disrupting chemicals (EDCs). EDCs are commonly found in natural and industrial sources and represent a large and growing family of compounds and mixtures with high chemical diversity. The Texas A&M Superfund Research Center is focused on evaluating and reducing the human health risks posed by exposure to hazardous substances, including EDCs, that are mobilized from sediments and other sources by environmental disaster events such as tropical storms, hurricanes, and floods. The overarching goal of Project 4 is to develop fast, robust, and cost-effective assays to determine the endocrine disrupting potential of complex, environmentally relevant chemical mixtures in the highly industrial Galveston Bay/Houston Ship Channel area (GB/HSC) and other Superfund sites. The availability of a set of fast, sensitive, and reproducible EDC assays will facilitate health hazard evaluation and improve decision-making in response to environmental emergencies. The central hypothesis that will be tested in Project 4 is that multi-parametric, highly mechanistic contextual in vitro assays, and bioinformatics analyses will serve as a robust, fast, and cost- effective framework to evaluate the endocrine disrupting potential of environmentally-derived complex chemical mixtures. Nuclear Receptors (NRs), a large class of transcription factors, are key mechanistic targets of EDCs. The assays developed in Project 4 will focus on three NRs identified by the Endocrine Disruptor Screening and Testing Advisory Committee: estrogen receptor (ER), androgen receptor (AR), and thyroid hormone receptor (TR). We will use advanced high throughput imaging and image analysis, genome-wide epigenomics, and integrative bioinformatics to address critical needs in assessing the risk to human health posed by hazardous substances: 1) Development of fast, robust, and cost effective high throughput assays to identify the presence of EDCs and classify their bioactivity. Cell-based platforms in endocrine-relevant systems will be used to analyze both single compounds (~50, including heavy metals, pesticides, pharmaceuticals, etc.), and complex mixtures, including samples from Superfund sites. 2) Comprehensive analysis of EDC impact on the epigenome and the identification of EDC-specific epigenetic ?fingerprints.? The epigenetic impact of EDCs has not previously been considered as a way to identify and classify EDCs, and has the potential to provide new insights into EDC- and NR-specific mechanisms of pathway disruption. 3) Assessment of the endocrine disrupting potential of chemical mixtures in the environment. While previous work has primarily focused on EDC activity of single compounds mediated by the ER, we will increase the scope and relevance of EDC assessment by measuring effects on AR and TR, and by moving from single compounds to mixtures. High content, cell-based, and epigenomic data will be translated into a bioinformatics-based framework to determine and predict the endocrine disrupting potential of complex, environmentally relevant chemical mixtures.